The bacteria Klebsiella oxytoca, isolated from the interior of Coffea arabica beans, obtained from the Universidade Federal de Viçosa plantation, in Viçosa, MG, was grown in 2.8 L Erlenmeyer flasks containing 1,0 L of minimal medium, pH 7,0, and Sigma citric pectin as sole carbon source, at a final concentration of 0.3% (w/ v). The culture was incubated at 25ºC, without shaking,
for 24 hours and reached a population of 108 cells per milliliter in less than 20 hours. Generation time under these conditions corresponded to 60 minutes. The growth rate at 37ºC, in this medium, differed little from the rates obtained at 25ºC and 30ºC. K. oxytoca proved capable of growing in this same medium without ammonium sulfate even under anaerobic conditions. The pectinolytic enzyme produced was induced intracellular pectate lyase, which acts primarily on unmethylated substrate. Optimal activity of this enzyme was obtained at a temperature of 40ºC, pH 9.0, in the presence of magnesium ions. The pectate lyase
presented a Km of 4.51 mg/ mL and did not lose activity when stored at -18ºC for 9 months or when held at 40ºC for 1 hour and 40 minutes. The physico-chemical characteristics of the pectinase produced were similar to those found in other bacteria and fungi. The bacteria secreted pectate lyase when grown in medium containing pectin of molecular mass greater than 30,000 Da, obtained by ultrafiltration. The enzyme's secretion was inhibited by a compound present in the pectin of molecular mass between 3,000 e 30,000 Da or by oligogalacturonides, which could somehow be degraded by the bacteria. The substrate retained inside
2,000 and 14,000 Da dialysis membranes induced greater enzyme secretion, being greatest in the 14,000 Da membranes. The compounds present in the Sigma citric pectin that interfered in enzyme regulation mechanisms were not identified. K. oxytoca quickly grewn when starch, galacturonate, poligalacturonate, casein, and glucose were used as carbon source. However, growth was not observed when using cellulose as carbon source. The depolymerizing enzymes were detected at minimal levels, a characteristic of endophytic microorganisms. Endophytic bacteria, that don’t make visible damage in the plants, should have some regulatory mechanism to control the synthesis and the activity of their enzymes. The results indicate the potential contribution of K. oxytoca in fermentative processes in coffee beans.